Interpretive Summary: Marek’s disease virus (MDV) is a serious pathogen for chickens, so understanding how this virus causes disease may provide valuable clues for improved control measures. MDV is highly cell associated, which poses problems for scientific investigations. To alleviate this hindrance, we generated a recombinant virus that fluoresces. Thus, this modification to the virus allows scientists to track the virus in living cells. Use of this model will provide critical information on how the virus replicates, spreads, and potentially causes disease in chickens.

Technical Abstract:
Marek’s disease virus (MDV) is a strictly cell-associated avian alphaherpesvirus. Although its replication was supported in chicken embryo fibroblasts (CEF) or duck embryo fibroblasts, identification of MDV-infected cell is quite cumbersome especially in the early stage of virus replication. To visualize MDV replication in infected cells and characterize MDV US10 in vitro, a recombinant MDV, rMd5-US10-EGFP virus, was generated that expresses enhanced green fluorescent protein (EGFP) as a tagged protein fused with US10 at the C-terminal end. The expression of US10-EGFP was detected in infected CEF using fluorescent microscopy and the expression intensity was quantified using flow cytometry analysis. In addition, confocal microscopic analysis provided information on sub-cellular localization of US10-EGFP in virus-infected cells, which is exclusively localized in the cytoplasm. In conclusion, rMd5-US10-EGFP virus can be used to test the early stage of infected cell, as well as to real-time monitor virus activity in vitro.